Structure and Plasticity of Silent Synapses in Developing

Xu et al. Cell Discovery (2020) 6:8 Cell Discovery https://doi.org/10.1038/s41421-019-0139-1 www.nature.com/celldisc

ARTICLE Open Access Structure and plasticity of silent synapses in developing hippocampal neurons visualized by super-resolution imaging Cheng Xu1,2, Hui-Jing Liu2,3,LeiQi2,3, Chang-Lu Tao1,Yu-JianWang1,ZeyuShen2, Chong-Li Tian2,3, Pak-Ming Lau2,3 and Guo-Qiang Bi1,2,4

Abstract Excitatory synapses in the mammalian brain exhibit diverse functional properties in transmission and plasticity. Directly visualizing the structural correlates of such functional heterogeneity is often hindered by the diffraction-limited resolution of conventional optical imaging techniques. Here, we used super-resolution stochastic optical reconstruction microscopy (STORM) to resolve structurally distinct excitatory synapses formed on dendritic shafts and spines. The majority of these shaft synapses contained N-methyl-D-aspartate receptors (NMDARs) but not α-amino-3- hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs), suggesting that they were functionally silent. During development, as more spine synapses formed with increasing sizes and expression of AMPARs and NMDARs, shaft synapses exhibited moderate reduction in density with largely unchanged sizes and receptor expression. Furthermore, upon glycine stimulation to induce chemical long-term potentiation (cLTP), the previously silent shaft synapses became functional shaft synapses by recruiting more AMPARs than did spine synapses. Thus, silent shaft synapse may represent a synaptic state in developing neurons with enhanced capacity of activity-dependent potentiation. 1234567890():,; 1234567890():,; 1234567890():,; 1234567890():,;

– Introduction functional form through activity-dependent plasticity13 16. In the mammalian brain, excitatory communication However, the structural and morphological correlates of between neurons is primarily mediated by glutamatergic these functional states have been lacking. Studies with synapses1,2. Activity-induced plasticity of these synapses is electron microscopy (EM) have indicated that most glu- believed to underlie learning and memory function of the tamatergic excitatory synapses are formed on dendritic – brain3 6. Electrophysiological studies have suggested that spines, in contrast to GABAergic inhibitory synapses that excitatory synapses may exhibit distinct functional prop- are primarily formed on dendritic shafts, although erties or states7,8. An extreme case is the so-called silent exceptions have been observed that some excitatory – – synapse9 12, which contains few α-amino-3-hydroxy-5- synapses formed directly on the shafts17 20. With con- methyl-4-isoxazolepropionic acid receptors (AMPARs) ventional ﬂuorescence microscopy, it was observed that and cannot carry out excitatory transmission upon pre- early in development, N-Methyl-D-aspartate receptors synaptic activation, but can be converted into the (NMDARs) clusters might form on dendritic shafts before clustering of AMPARs21. Unfortunately, the diffraction- limited resolution of conventional optical microscopy Correspondence: Pak-Ming Lau ([email protected])or does not allow for unambiguous determination whether Guo-Qiang Bi ([email protected]) 1Hefei National Laboratory for Physical Sciences at the Microscale, University of these receptor clusters are actual shaft synapses. Thus, a Science and Technology of China, Hefei, Anhui 230027, China higher-resolution imaging approach is desired to establish 2 School of Life Sciences, University of Science and Technology of China, Hefei, the link between the morphological and functional states Anhui 230027, China Full list of author information is available at the end of the article. of these synapses. In the current study, we took advantage These authors contributed equally: Cheng Xu, Hui-Jing Liu